Heart failure is major cause of mortality associated with mostly Myocardial infarction (MI). Transplanting mesenchymal stem cells (MSC) have exhibited potential role in myocardial regeneration. Secretion of immune-modulatory cytokines and various growth factors after transplantation plays significant role in remodelling process of MI region. However, low retention, higher shear stress during administration and rejection at host infarct environment hinders therapeutic efficacy. Myocardial regeneration demands for accurate spatio-temporal delivery of MSCs with supportive vascular network that leads to improvement of cardiac function. In this study, injectable alginate based microporous hydrogel has been used to deliver 5-Azacytidine (5-Aza) in zein protein nanoparticle with MSCs for attenuating adverse cardiac remodelling after MI. Zein nanoparticles loaded with 5-Aza were prepared by liquid–liquid dispersion, and it was found that 35% of drug was released in 7 days supported with mathematical modelling. The presence of 5-Aza and zein in developed hydrogel supported in vitro MSC proliferation, migration and angiogenesis. Significant increased expression of cardiac specific markers, GATA4, MEF2C, MLC, SERCA and NKX2.5 was observed in vitro. 5-Aza loaded protein nanoparticle with MSCs encapsulated hydrogels in rat MI model also exhibited substantial improvement of functional cardiac parameters such as cardiac output and ejection fraction. Histopathological analysis showed reduced fibrosis, attenuated infarct expansion and cardiac tissue restoration and angiogenesis. In brief, we developed nanocarrier-hydrogel system a promising strategy for co-delivering 5-Aza as cardiac differentiation cue with MSCs to achieve higher cell retention and enhanced improvement in myocardial regeneration after MI. © 2021 Elsevier B.V.